The present invention relates to a fast method for the determination of a sequence of a nucleic acid, DNA or RNA, which is useful, in particular, for the sequencing of an unknown DNA or RNA or alternatively for the detection of a specific DNA or RNA sequence for diagnosis.
DNA sequencing is a major objective of molecular biology, and is at the heart of the human genome sequencing project.
Furthermore, demonstration of the presence of a specific DNA sequence in a physiological sample constitutes, at the present time, the major line of development of diagnostic methods. After the immunological technique, attention is now being turned to diagnostic methods that demonstrate modifications of the DNA itself in order to prevent, in particular, antibiotic resistance (see Nature, 1992, 358, p. 591), genetic abnormalities, the risks of cancer associated with genetic modifications and viral infections, for example infections associated with HIV or with hepatitis viruses.
These diagnostic methods comprise, at the present time, the so-called "direct hybridization" methods, which will detect the presence of this sequence by direct hybridization of a probe with the sample. The method is cumbersome and imprecise, in particular because it necessitates a detection by gel and exposure on light-sensitive film or radioactive counting.
Methods employing an amplification, in particular the PCR method, which, before the demonstration of the sequence by hybridization, will first amplify the corresponding portion of the DNA using, for example, primers and a polymerase, are excessively sensitive to contamination.
These methods have, in addition, the drawback of being indirect recognition systems, because the desired sequence is recognized only via the use of A probe.